COVID-19 was associated with the complications after allogeneic hematopoietic stem cell transplantation

We aimed to identify the severity and duration of COVID-19 infection on complications after allo-HSCT. Enrolled 179 hospitalized patients with COVID-19 were categorized into long-term infection (> 18 days, n = 90) or short-term infection group (≤ 18 days, n = 89) according to the median duration of COVID-19. The severity of COVID-19 was categorized as asymptomatic infection, mild, moderate, severe, and critical illness according to guidelines of National Institutes of Health. Particularly, severe illness and critical illness were classified as serious infection. Asymptomatic infection, mild illness and moderate illness were classified as non-serious infection. The 150-day probabilities of poor graft function (PGF), cytomegalovirus (CMV) pneumonia and non-relapse mortality (NRM) were significantly higher in long-term infection group. The 150-day probabilities of CMV pneumonia and NRM after COVID-19 were higher in serious infection group. The 150-day probabilities of overall survival (OS) was significantly lower in long-term and serious infection group. In multivariable analysis, the severity of COVID-19 was associated with NRM and OS, and the duration of COVID-19 was associated with PGF. In summary, our data reported that the severity and duration of COVID-19 were associated with several complications and contribute to poor outcomes after allo-HSCT.

www.nature.com/scientificreports/COVID-19 before 2021 which might be not the same as the Omicron variants.In the Omicron era, several studies have identified the characteristics of COVID-19 in immunocompromised patients, but most of them also only focused on the clinical presentations and treatments of COVID-19, and the sample of allo-HSCT recipients was small 7,8,[20][21][22] .Thus far, no study had focused on the association between occurrence, severity, and duration of COVID-19 infection and complications after allo-HSCT.Particularly, the influence of Omicron variant infection on the post-transplant complications is unclear.
Thus, we aimed to identify the severity and duration of COVID-19 on post-transplant complications in the Omicron era.
The median duration of leukopenia, thrombocytopenia, and PGF was 14 days (range 4-118) days, 10 days (range 8-78) days, and 17 days (range 9-78) days, respectively.Until the last follow up, leukopenia, thrombocytopenia and PGF were still persistence 22 patients.2).The other risk factors for PGF and leukopenia were showed in Supplementary Table 4.

Discussion
In the present study enrolled consecutive hospitalized COVID-19 patients, 24% of them showed serious infection, and more than half of the patients experienced long-term COVID-19 infection (i.e., more than 18 days).The 150-day cumulative incidence of PGF, leukopenia, CMV disease, CMV pneumonia, NRM, and OS after COVID-19 was significantly higher in long-term infection group compared with short-term infection group.In addition, the 150-day cumulative incidence of CMV disease, CMV pneumonia, NRM, and OS after COVID-19 was also significantly higher in serious infection group compared with non-serious infection group.Thus far, this is the first study identify the influence of severity and duration of SARS-CoV-2 infection on complications after allo-HSCT in the Omicron era.Viral infection is one of the most important causes of PGF.Parvovirus B19 was the most common virus which can lead to rejection and PGF 24,25 .In addition, herpes virus could also induce PGF.For example, the association of HHV-6 infections and thrombocytopenia has been reported since 1990s 26 , and several studies observed the association between HHV-6 infections and delayed platelet engraftment in allo-HSCT recipients 27,28 .Some studies also reported that CMV infection was associated with thrombocytopenia 29 and PGF 16 .The potential mechanism including impairment of virus on hematopoietic microenvironment or hematologic toxicities of anti-virus drugs.Herpes simplex virus and CMV can infect hematopoietic progenitor cells [30][31][32] , and the treatments of CMV such as gancilovir and forscarnet could lead to myelosuppression.Similarly, several studies also reported SARS-CoV-2 infection could cause anemia [33][34][35] , thrombocytopenia [34][35][36] , and aplastic anemia 37 .Thus, it is suggested that we should pay attention to PGF in patients with SARS-CoV-2 infection, particularly for those with long-term infection.
In addition, we observed that the incidence of CMV disease, particularly the CMV pneumonia, significantly increased in those with serious and/or long-term SARS-CoV-2 infection.Some data suggested that some viruses, such as HHV-6 and HHV-7, could take part in CMV reactivation or progression in immunosuppressed patients 38,39 .Lymphopenia is the hallmark of severe COVID-19 presentation, and previous studies observed that both the numbers of circulating T, B, and NK cells and antiviral cytokine production capability decreased in patients with COVID-19, particularly in those required intensive care [40][41][42] .Drylewicz et al. 43 show that reconstitution of CD4 + T cells and NK cells after allo-HSCT is important for CMV prophylaxis.Thus, we speculated that COVID-19 infection may increase the susceptibility of CMV reactivation after allo-HSCT, which should be further confirmed in the future.
Previous studies have shown that the mortality rate range 12.5-22% in allo-HSCT recipients after COVID-19 infection [44][45][46] .While in the immunodeficient patients infected with Omicron variant, the mortality rate was 10.5% in the advanced malignancy cohort 23 .Our data suggested the 150-day cumulative incidence of NRM after COVID-19 infection was 11.2% in allo-HSCT recipients, which was similar to the previous studies.In addition, we only enrolled the hospitalized COVID-19 patients, which may also contribute to the relatively high NRM.
As mentioned above, long-term SARS-CoV-2 infection increased the risk of PGF, CMV disease, and NRM, this suggested that how to clear the SARS-CoV-2 timely is critical to improve the outcomes of these patients.Besides of the anti-viral drug (e.g., Nirmatrelvir/Ritonavir), the intense of immunosuppressive therapy could influence the viral clearance and was associated with severity and persistence of infection.Thus, allo-HSCT recipients may taper the immunosuppressive therapy after SARS-CoV-2 infection.
There were some limitations in the present study.This was a single center, observational cohort study and the followed-up was still relatively short, so we should further identify the long-term impact of Omicron variant infection on post-transplant complications in future.
In summary, we observed that the severity and duration of COVID-19 infection were associated with PGF, leukopenia, CMV disease, CMV pneumonia, NRM, and survival after allo-HSCT.Thus, how to prevent and clear the SARS-CoV-2 infection effectively is still critical for allo-HSCT recipients in the Omicron era.

Patients
We performed an observational prospective study in Peking University, Institute of Hematology (PUIH, n = 179).Including criteria were as followed: 1) the hospitalized patients had received an allo-HSCT in PUIH at any time before the diagnosis of COVID-19, and 2) have laboratory-confirmed (polymerase chain reaction or antigenic test) COVID-19.In order to compare the clinical outcome between patients with and without COVID-19, a historical cohort included patients receiving allo-HSCT from August 1 2016 to December 1 2022 were enrolled and they were matched for age, sex, underlying disease, allo-HSCT time and time after allo-HSCT in a 1: 1 ratio.The study was approved by the Ethics Committee of Peking University People's Hospital, and written informed consent was obtained from all subjects before study entry, in accordance with the Declaration of Helsinki.

Transplantation protocols
Donor selection, human leukocyte antigen typing, graft harvesting, conditioning regimen, GVHD, and infection prophylaxis were performed as previously described 1 .Comorbidities in HSCT recipients were assessed according to the hematopoietic cell transplantation-specific comorbidity index (HCT-CI).

Clinical definitions and assessments
During our study period, the Omicron variant was predominant in Beijing, China.The severity of COVID-19 was categorized as asymptomatic infection, mild, moderate, severe, and critical illness according to guidelines of National Institutes of Health (Supplementary methods).Severe illness and critical illness were classified as serious infection, and asymptomatic infection, mild illness and moderate illness were classified as non-serious infection in the present study.The duration of COVID-19 infection was defined as the time of the patients with persistent signs and symptoms with polymerase chain reaction or antigenic test positivity.
PGF was defined as the presence of 2 or 3 cytopenic counts (ANC ≤ 0.5 × 10 9 /L, platelet ≤ 20 × 10 9 /L, or hemoglobin ≤ 70 g/L) for at least 3 consecutive days beyond day 28 post-transplantation or dependence on transfusion and granulocyte colony-stimulating factor (G-CSF), in the presence of complete donor chimerism.Leukopenia was defined as ANC ≤ 0.5 × 10 9 /L alone for at least 3 consecutive days beyond day 28 post-transplantation or dependence on G-CSF, thrombocytopenia was defined as engraftment of peripheral blood cell lines (ANC > 0.5 × 10 9 /L and hemoglobin > 70 g/L without transfusion support) other than a platelet count < 20 × 10 9 /L or dependence on platelet transfusions for at least 7 consecutive days, in the presence of complete donor chimerism.Patients with evidence of hematological relapse post-HSCT were excluded.GVHD was diagnosed and graded based on international criteria 47 .CMV infection was diagnosed according to the definition of CMV Drug Development Forum 48 .PTLD was diagnosed according to the Sixth European Conference on Infections in Leukemia guidelines 49 .
Relapse was defined by morphologic evidence of disease in peripheral blood, bone marrow, or extramedullary site samples or by the recurrence and sustained presence of pre-transplantation chromosomal abnormalities.www.nature.com/scientificreports/NRM was defined as death without disease progression or relapse.OS was defined as the time from transplantation to death from any cause.

Statistical analysis
Characteristics of patients were summarized by descriptive statistics, that is, using counts and percentages for categorical variables and using median and range for continuous variables.Subject variables were compared using the χ 2 test for categorical variables and the Mann-Whitney U test for continuous variables.Multivariate analyses were performed using the Cox proportional hazards model for survival to identify the independent prognostic variables (Supplementary methods).The parameters with P < 0.10 according to the univariate analysis were entered into a multivariate model (Supplementary Table 3).Cumulative incidence curves were used in a competing risk setting, with relapse treated as a competing event, to calculate NRM probabilities, and with death and relapse as the competing risks for PGF, infection, and GVHD.The probability of survival was estimated with the Kaplan-Meier method and were compared using the log-rank test.Statistical analyses were performed using 1-way analysis of variance (ANOVA) for comparisons among the groups.Analyses were performed using GraphPad Prism 6.0 (GraphPad Software), SPSS 24 (SPSS Inc./IBM, Armonk, NY, USA) and R version 3.4.4(The R Foundation for Statistical Computing).Unless otherwise specified, all P values were 2-sided and P < 0.05 was considered significant.

Figure 2 .
Figure 2. The association between COVID-19 and CMV disease.The 150-day cumulative incidence of CMV disease after COVID-19 between (A) non-serious and serious infection group; (B) short and long-term infection group.The 150-day cumulative incidence of CMV pneumonia after COVID-19 between (C) nonserious and serious infection group; (D) short and long-term infection group.CMV cytomegalovirus.

Figure 3 .
Figure 3.The association between COVID-19 and survival.The 150-day cumulative incidence of NRM after COVID-19 between (A) non-serious and serious infection group; (B) short and long-term infection group.The 150-day probability of OS after COVID-19 infection between (C) non-serious and serious infection group; (D) short and long-term infection group.NRM non-relapse mortality, OS overall survival.

Figure 4 .
Figure 4. Clinical outcomes of patients with and without COVID 19 infection.(A) The 150-day cumulative incidence of NRM in the group with and without COVID-19 infection; (B) The 150-day cumulative incidence of OS in the group with and without COVID-19 infection; (C) The 150-day cumulative incidence of NRM in the group without COVID-19 infection, serious infection and long-term infection; (D) The 150-day cumulative incidence of OS in the group without COVID-19 infection, serious infection and long-term infection, NRM non-relapse mortality, OS overall survival. https://doi.org/10.1038/s41598-024-62731-7 Azifudine).Sixty-two patients received newly added drugs corticosteroid treatment, that is, 23 (53.5%) and 39 (28.7%) patients were in the serious and non-serious infection group, 9 (10.1%) and 53 (58.9%) patients were in the short-and long-term infection group, respectively.

Table 2 .
Multivariate analysis of risk factors for the 150-day clinical outcomes after COVID-19 infection.PGF poor graft function, CMV cytomegalovirus, NRM non-relapse mortality, OS overall survival.